Separate to operate: control of centrosome positioning and separation

Abstract

The centrosome is the main microtubule (MT)-organizing centre of animal cells. It consists of two centrioles and a multi-layered proteinaceous structure that surrounds the centrioles, the so-called pericentriolar material. Centrosomes promote de novo assembly of MTs and thus play important roles in Golgi organization, cell polarity, cell motility and the organization of the mitotic spindle. To execute these functions, centrosomes have to adopt particular cellular positions. Actin and MT networks and the association of the centrosomes to the nuclear envelope define the correct positioning of the centrosomes. Another important feature of centrosomes is the centrosomal linker that connects the two centrosomes. The centrosome linker assembles in late mitosis/G1 simultaneously with centriole disengagement and is dissolved before or at the beginning of mitosis. Linker dissolution is important for mitotic spindle formation, and its cell cycle timing has profound influences on the execution of mitosis and proficiency of chromosome segregation. In this review, we will focus on the mechanisms of centrosome positioning and separation, and describe their functions and mechanisms in the light of recent findings.

Keywords: Eg5; centrosome positioning; centrosome separation.

Figure 1.

The centrosome cycle with special focus on centriole disengagement. The centrosome cycle starts in early S-phase when the daughter centriole assembles next to the mother centriole. Short daughter centrioles are formed in early S-phase and further elongate in G2. With cytokinesis each daughter cell inherits one centrosome with two joint centrioles. In late mitosis/G1, the two centrioles separate from each other. This process is called ‘centriole disengagement’ (depicted in the enlargement at the right) and is regulated by the protease separase. Cleavage of the cohesin subunit Rad21^Scc1 and kendrin stimulates centriole separation [11,12]. It is unclear whether separase is regulated similarly at the centrosome and centromere. Securin and cyclin B1 are inhibitors of separase. At centrosomes a splice variant of Sgo1, sSgo1, additionally protects premature centriole disengagement. Moreover, Aki1 and astrin have been implicated as centrosome specific inhibitors of separase [13,14].

Figure 2.

Centrosome positioning around the nucleus. (a) In G2 phase, the mother centrosome is attached to the NPC. Two mechanisms account for this attachment. First, Nup358, a component of the NPC, minus-end directed motor protein dynein and the adaptor protein BICD2 form a complex which attaches to astral MTs emanating from the mother centrosome. Second, the N-terminal domain of Nup133 interacts with CENP-F that becomes positioned at the NE in G2/M. Dynein binds via the NudE/NudEL/CENP-F complex. Recruitment of BICD2 or CENP-F to the NPC does not depend on each other, which implies that the two pathways are independent from each other [45]. (b) When both centrosomes attach to the nucleus, the pull-and-push forces created by dynein and the kinesin motor Kif1 separate the centrosomes, and the centrosomes start to slide on the nucleus by being attached to the next NPC. (c) When they are separated enough to form anti-parallel MTs, dynein and the plus-end directed motor protein Eg5 become involved in further separation of the centrosomes. Dynein and Eg5 create opposing forces in order to balance the separation. Eg5 pushes the centrosomes away from each other, although dynein tries to keep them together by forming a complex with plus-end protein CLIP-170 and adaptor protein Lis1. Dynein localizes to MTs arising from one centrosome and catches the plus-end of MTs from the other centrosome. By pulling the MTs, dynein creates an inward force opposite to Eg5. (d) Prophase centrosome separation is completed when the separated centrosomes detach from the nucleus.

Figure 3.

Molecules involved in centrosome separation. Aurora A is found on the top of the linker dissolution pathway. Plk1 is activated upon phosphorylation by aurora A. Plk1, in turn, phosphorylates Mst2 that is in complex with hSav1. Phosphorylation of Mst2 by Plk1 increases the activity of Nek2A and decreases the activity of PP1γ towards C-Nap1 and rootletin, by altering the complex formation between these proteins [58]. Phosphorylated C-Nap1 and rootletin diffuse away from the centrosome, and the centrosomes become ready for the separation. Plk1 additionally promotes centrosome separation directly by phosphorylating Eg5 and indirectly by regulating the Nek9-Nek6/7-Eg5 pathway. Nek9 is also regulated by LC8, which is a part of dynein complex and functions in linking dynein to different cargo proteins [59]. LC8 enhances the activation of auto-phosphorylated Nek9. Moreover, binding of LC8 impairs the interaction between Nek9 and Nek6 [60]. Cdk1 is also involved in centrosome separation by directly phosphorylating Eg5 [61,62].